10/08/1402
The first report on isolation of a new highly hemolytic toxin, Scatotoxin, from Scatophagus argus venomous bonny spines
Scatophagus argus
of family Scatophagidae is a venomous fish. Rough spines insulated from
scat possessed potent venom composed of several proteins. Envenomation
is associated with local necrosis and severe pain. Following our
previous report regarding the hemolytic activity of scat crude venom,
this work aimed at purification and evaluation of its hemolytic protein,
hereafter designated as Scatotoxin. Specimens were collected from
coastal waters of the Persian Gulf, Iran. Proteins were extracted from
bone tissue by solubilization buffers and subsequently refolded in a
refolding buffer. Purification was performed by reverse-phase HPLC
method using a linear gradient protocol. To evaluate the hemolytic
activity of Scatotoxin, a quantitative microscopic assay was developed
using cell counting by which measurement of activity of the least amount
of a sample was achievable. Scatotoxin was isolated in 85%
acetonitrile. It is an interesting highly hydrophobic protein. Because
hemolysis was observed immediately, scatotoxin is considered a very
fast-acting hemolytic agent. Scatotoxin indicated as a 72 kDa protein by
SDS-PAGE. The amount of 0.5µg crude venom produced 100% hemolysis and
HD50 determined at 0.18µg. HD50 for scatotoxin recorded at 0.003µg. High
efficiency of both extraction method and microscopic-scale assay led to
the reduction of collected specimens and consequently avoiding harmful
effects on the Persian Gulf ecosystem. This issue is ethically important
due to decreasing the number of samples too. Among the previously
reported hemolytic proteins, Scatotoxin is the first report of a highly
hydrophobic protein.